Hönscheid, Pia

Abstract Neuropilin-2 (NRP2) is a non-tyrosine kinase receptor frequently overexpressed in various malignancies, where it has been implicated in promoting many protumorigenic behaviors, such as imparting therapeutic resistance to metastatic cancer cells. Here, we report a novel function of NRP2 as a regulator of endocytosis, which is enhanced in cancer cells and is often associated with increased metastatic potential and drug resistance. We found that NRP2 depletion in human prostate and pancreatic cancer cells resulted in the accumulation of EEA1/Rab5-positive early endosomes concomitant with a decrease in Rab7-positive late endosomes, suggesting a delay in early-to-late endosome maturation. NRP2 depletion also impaired the endocytic transport of cell surface EGFR, arresting functionally active EGFR in endocytic vesicles that consequently led to aberrant ERK activation and cell death. Mechanistic investigations revealed that WD-repeat– and FYVE-domain–containing protein 1 (WDFY1) functioned downstream of NRP2 to promote endosome maturation, thereby influencing the endosomal trafficking of EGFR and the formation of autolysosomes responsible for the degradation of internalized cargo. Overall, our results indicate that the NRP2/WDFY1 axis is required for maintaining endocytic activity in cancer cells, which supports their oncogenic activities and confers drug resistance. Therefore, therapeutically targeting endocytosis may represent an attractive strategy to selectively target cancer cells in multiple malignancies. Cancer Res; 76(2); 418–28. ©2015 AACR.

Abstract Radiotherapy is an important treatment regimen for prostate cancer. We have shown that the lymphangiogenic growth factor, Vascular Endothelial Growth Factor C (VEGF-C), induces docetaxel resistance and autophagy in prostate cancer (Muders et al., Cancer Res 2009 and Stanton et al., Cancer Res 2013). In this study we have evaluated the role of VEGF-C and VEGF-C induced autophagy in radioresistance of prostate cancer using cell culture experiments and human tissue samples. The human prostate carcinoma cell lines PC-3, LNCaP and DU145 were used in these cell culture experiments. Clonogenic survival of prostate cancer cells, after RNA interference (RNAi) of VEGF-C, autophagy related gene 5 (ATG5), and after addition of recombinant human VEGF-C following irradiation therapy, was evaluated. Autophagic flux was tested after ionizing irradiation using a Light-Chain-3 II immunoblot. To confirm our in vitro findings, human tissues of patients who underwent radical prostatectomy and adjuvant radiotherapy at Mayo Clinic were evaluated for VEGF-C expression. The quantity and intensity of VEGF-C staining was evaluated by two pathologists independently and correlated with biochemical relapse free survival (BRFS). The follow-up time is up to 23 years after radiotherapy. Multivariate Cox analysis was performed. VEGF-C levels correlated with a significantly higher radioresistance in colony formation assays and human tissue samples by multivariate analysis. The risk of biochemical recurrence increased 2.8 fold (95% confidence interval: 1.109 to 7.237; p = 0.03) when VEGF-C was highly expressed in prostatectomy patients with adjuvant radiotherapy. In line with our studies on autophagy in VEGF-C mediated docetaxel resistance of prostate cancer, autophagic flux was reduced after VEGF-C depletion during radiation. Interestingly, RNAi for ATG5 showed no effect on radiosensitivity which suggests that the decrease in autophagic flux after VEGF-C depletion is not responsible for the increased radiosensitivity. We conclude that VEGF-C is an important mediator of radiotherapy resistance in prostate cancer. In contrast to our published results on chemotherapy resistance, VEGF-C induced autophagy appears to be less important for resistance against ionizing radiation. Other VEGF-C dependent mechanisms for therapy resistance are under investigation. Citation Format: Steffi Haberlau, Pia Hönscheid, Rafael E. Jimenez, Gustavo B. Baretton, Donald J. Tindall, Mechthild Krause, Kaustubh Datta, Michael H. Muders. Role of Vascular Endothelial Growth Factor C in promoting radioresistance of prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3304. doi:10.1158/1538-7445.AM2015-3304

Abstract A major contributor to cancer mortality is recurrence and subsequent metastatic transformation following therapeutic intervention. Therefore, in order to develop new treatment modalities and improve the efficacy of current ones, it is important to understand the molecular mechanisms that promote resistance to therapy in cancer cells. One pathway contributing to therapy resistance is autophagy, a self-digestive process that can eliminate unnecessary or damaged organelles to protect cancer cells from death. We have found that the VEGF-C/NRP-2 axis is involved in the activation of autophagy, which helps cancer cell survival following treatment. Inhibition of mTOR complex 1 activity by this axis is the underlying mechanism for the activation of autophagy. Furthermore, we identified two VEGF-C/NRP-2-regulated genes, LAMP-2 and WDFY-1, that have previously been suggested to participate in autophagy and vesicular trafficking. Upregulation of WDFY-1 following VEGF-C or NRP-2 depletion contributes to cytotoxic drug-mediated cell death. Together, these data suggest a link between the VEGF-C/NRP-2 axis and cancer cell survival despite the presence of chemotherapy-induced stress. Effective targeting of this pathway may lead to the development of new cancer therapies. Cancer Res; 73(1); 160–71. ©2012 AACR.